PKC‐ζ is required for angiotensin II‐induced activation of ERK and synthesis of C‐FOS in MCF‐7 cells

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Abstract
We examined the signalling pathways responsible for the Ang II induction of growth in MCF‐7 human breast cancer cells. Ang II in MCF‐7 cells induced: (a) the translocation from the cytosol to membrane and nucleus of atypical protein kinase C‐ζ (PKC‐ζ) but not of PKC‐α, ‐δ, ‐ε and ‐η; (b) the expression of c‐fos mRNA and protein; (c) the phosphorylation of the extracellular signal‐regulated protein kinases 1 and 2 (ERK1/2). All these effects were due to the activation of the Ang II type I receptor (AT1) since they were blocked by the AT1 antagonist losartan. The Ang II‐stimulated ERK1/2 phosphorylation was blocked by (a) high doses of staurosporine, inhibitor of PKC‐ζ, and by a synthetic myristoylated peptide with sequences based on the endogenous PKC‐ζ pseudosubstrate region (ζ‐PS); (b) PD098059, a mitogen‐activated protein kinase kinase inhibitor (MAPKK/MEK); and, moreover, (c) the inhibitors of phosphoinositide 3‐kinases (PI3K), LY294002 and wortmannin, thus indicating that PI3K may act upstream of ERK1/2. The Ang II‐evoked c‐fos induction was blocked only by high doses of staurosporine and by ζ‐PS whilst PD098059, LY294002 and wortmannin were ineffective, thus indicating that c‐fos induction is not due to ERK1/2 activity. When the epidermal growth factor‐receptor (EGFR) tyrosine kinase activity was inhibited by the use of its inhibitor AG1478, Ang II was still able to induce ERK1/2 phosphorylation and c‐fos expression, therefore proving that the transactivation of EGFR was not required for these Ang II effects in MCF‐7 cells. The previously reported proliferation of MCF‐7 cells induced by Ang II was blocked by PD098059 and by wortmannin in a dose‐dependent manner, thereby indicating that in MCF‐7 cells the PI3K and ERK pathways mediate the mitogenic signalling of AT1. Our results suggest that in MCF‐7 cells Ang II activates multiple signalling pathways involving PKC‐ζ, PI3K and MAPK; of these pathways only PKC‐ζ appears responsible for the induction of c‐fos. J. Cell. Physiol. 197: 61–68, 2003